Regional synchrony in full-scale activated sludge bioreactors due to deterministic microbial community assembly.
ABSTRACT: Seasonal community structure and regionally synchronous population dynamics have been observed in natural microbial ecosystems, but have not been well documented in wastewater treatment bioreactors. Few studies of community dynamics in full-scale activated sludge systems facing similar meteorological conditions have been done to compare the importance of deterministic and neutral community assembly mechanisms. We subjected weekly activated sludge samples from six regional full-scale bioreactors at four wastewater treatment plants obtained over 1 year to Illumina sequencing of 16S ribosomal RNA genes, resulting in a library of over 17 million sequences. All samples derived from reactors treating primarily municipal wastewater. Despite variation in operational characteristics and location, communities displayed temporal synchrony at the individual operational taxonomic unit (OTU), broad phylogenetic affiliation and community-wide scale. Bioreactor communities were dominated by 134 abundant and highly regionally synchronized OTU populations that accounted for over 50% of the total reads. Non-core OTUs displayed abundance-dependent population synchrony. Alpha diversity varied by reactor, but showed a highly reproducible and synchronous seasonal fluctuation. Community similarity was dominated by seasonal changes, but individual reactors maintained minor stable differences after 1 year. Finally, the impacts of mass migration driven by direct biomass transfers between reactors was investigated, but had no significant effect on community similarity or diversity in the sink community. Our results show that population dynamics in activated sludge bioreactors are consistent with niche-driven assembly guided by seasonal temperature fluctuations.
Project description:Denitrifying phosphorus removal is an attractive wastewater treatment process due to its reduced carbon source demand and sludge minimization potential. Two lab-scale sequencing batch reactors (SBRs) were operated in alternating anaerobic-anoxic (A-A) or anaerobic-oxic (A-O) conditions to achieve denitrifying enhanced biological phosphate removal (EBPR) and traditional EBPR. No significant differences were observed in phosphorus removal efficiencies between A-A SBR and A-O SBR, with phosphorus removal rates being 87.9% and 89.0% respectively. The community structures in denitrifying and traditional EBPR processes were evaluated by high-throughput sequencing of the PCR-amplified partial 16S rRNA genes from each sludge. The results obtained showed that the bacterial community was more diverse in A-O sludge than in A-A sludge. Taxonomy and ?-diversity analyses indicated that a significant shift occurred in the dominant microbial community in A-A sludge compared with the seed sludge during the whole acclimation phase, while a slight fluctuation was observed in the abundance of the major taxonomies in A-O sludge. One Dechloromonas-related OTU outside the 4 known Candidatus "Accumulibacter" clades was detected as the main OTU in A-A sludge at the stationary operation, while Candidatus "Accumulibacter" dominated in A-O sludge.
Project description:Nitrous oxide (N<sub>2</sub>O) is a strong greenhouse gas and causal for stratospheric ozone depletion. During biological nitrogen removal in wastewater treatment plants (WWTP), high N<sub>2</sub>O fluxes to the atmosphere can occur, typically exhibiting a seasonal emission pattern. Attempts to explain the peak emission phases in winter and spring using physico-chemical process data from WWTP were so far unsuccessful and new approaches are required. The complex and diverse microbial community of activated sludge used in biological treatment systems also exhibit substantial seasonal patterns. However, a potentially causal link between the seasonal patterns of microbial diversity and N<sub>2</sub>O emissions has not yet been investigated. Here we show that in a full-scale WWTP nitrification failure and N<sub>2</sub>O peak emissions, bad settleability of the activated sludge and a turbid effluent strongly correlate with a significant reduction in the microbial community diversity and shifts in community composition. During episodes of impaired performance, we observed a significant reduction in abundance for filamentous and nitrite oxidizing bacteria in all affected reactors. In some reactors that did not exhibit nitrification and settling failures, we observed a stable microbial community and no drastic loss of species. Standard engineering approaches to stabilize nitrification, such as increasing the aerobic sludge age and oxygen availability failed to improve the plant performance on this particular WWTP and replacing the activated sludge was the only measure applied by the operators to recover treatment performance in affected reactors. Our results demonstrate that disturbances of the sludge microbiome affect key structural and functional microbial groups, which lead to seasonal N<sub>2</sub>O emission patterns. To reduce N<sub>2</sub>O emissions from WWTP, it is therefore crucial to understand the drivers that lead to the microbial population dynamics in the activated sludge.
Project description:Wastewater treatment plants in temperate climate zones frequently undergo seasonal nitrification failure in the winter month yet maintain removal efficiency for other contaminants. We tested the hypothesis that nitrification failure can be correlated to shifts in the nitrifying microbial community. We monitored three parallel, full-scale sequencing batch reactors over the course of a year with respect to reactor performance, microbial community composition via 16S rRNA gene amplicon sequencing, and functional gene abundance using qPCR. All reactors demonstrated similar changes to their core microbiome, and only subtle variations among seasonal and transient taxa. We observed a decrease in species richness during the winter, with a slow recovery of the activated sludge community during spring. Despite the change in nitrification performance, ammonia monooxygenase gene abundances remained constant throughout the year, as did the relative sequence abundance of Nitrosomonadacae. This suggests that nitrification failure at colder temperatures might result from different reaction kinetics of nitrifying taxa, or that other organisms with strong seasonal shifts in population abundance, e.g. an uncultured lineage of Saprospiraceae, affect plant performance in the winter. This research is a comprehensive analysis of the seasonal microbial community dynamics in triplicate full-scale sequencing batch reactors and ultimately strengthens our basic understanding of the microbial ecology of activated sludge communities by revealing seasonal succession patterns of individual taxa that correlate with nutrient removal efficiency.
Project description:Conventional and advanced biological wastewater treatment systems generate excess sludge, which causes socio-economic and environmental issues. This study investigated the performance of membrane-controlled anoxic-oxic-anoxic (AOA) bioreactors for in-situ sludge reduction compared to the conventional anoxic-oxic-oxic membrane bioreactor (MBR<sub>control</sub>). The membrane units in the AOA bioreactors were operated as anoxic reactors at lower sludge recirculation rates to achieve hydrolysis of extracellular polymeric substances (EPS) and extensive endogenous respiration. Compared to MBR<sub>control</sub>, the AOA bioreactors operated with 90%, and 80% recirculation rates reduced the sludge growth up to 19% and 30%, respectively. Protein-like components were enriched in AOA bioreactors while fulvic-like components were dominant in MBR<sub>control</sub>. The growth of <i>Dechloromonas</i> and <i>Zoogloea</i> genra was promoted in AOA bioreactors and thus sludge reduction was facilitated. Metagenomics analysis uncovered that AOA bioreactors exhibited higher proportions of key genes encoding enzymes involved in the glycolysis and denitrification processes, which contributed to the utilization of carbon sources and nitrogen consumption and thus sludge reduction.
Project description:The development and activity of a cold-adapting microbial community was monitored during low-temperature anaerobic digestion (LtAD) treatment of wastewater. Two replicate hybrid anaerobic sludge bed-fixed-film reactors treated a synthetic sewage wastewater at 12°C, at organic loading rates of 0.25-1.0 kg chemical oxygen demand (COD) m-3 d-1, over 889 days. The inoculum was obtained from a full-scale anaerobic digestion reactor, which was operated at 37°C. Both LtAD reactors readily degraded the influent with COD removal efficiencies regularly exceeding 78% for both the total and soluble COD fractions. The biomass from both reactors was sampled temporally and tested for activity against hydrolytic and methanogenic substrates at 12°C and 37°C. Data indicated that significantly enhanced low-temperature hydrolytic and methanogenic activity developed in both systems. For example, the hydrolysis rate constant (k) at 12°C had increased 20-30-fold by comparison to the inoculum by day 500. Substrate affinity also increased for hydrolytic substrates at low temperature. Next generation sequencing demonstrated that a shift in a community structure occurred over the trial, involving a 1-log-fold change in 25 SEQS (OTU-free approach) from the inoculum. Microbial community structure changes and process performance were replicable in the LtAD reactors.
Project description:Understanding spatiotemporal patterns in microbial community composition is a central goal of microbial ecology. The objective of this study was to better understand the biogeography of activated sludge microbial communities, which are important for the protection of surface water quality. Monthly samples were collected from 20 facilities (25 bioreactors) within 442 km of each other for 1 year. Microbial community composition was characterized by sequencing of PCR-amplified 16S rRNA gene fragments. Statistically significant distance decay of community similarity was observed in these bioreactors independent of clustering method (operational taxonomic units [OTUs] at 97% similarity, genus-level phylotypes) and community dissimilarity metric (Sørensen, Bray-Curtis, and weighted Unifrac). Universal colonizers (i.e., detected in all samples) and ubiquitous genus-level phylotypes (i.e., detected in every facility at least once) also exhibited a significant distance decay relationship. Variation partitioning analysis of community composition showed that environmental characteristics (temperature, influent characteristics, etc.) explained more of the variance in community composition than geographic distance did, suggesting that environmental heterogeneity is more important than dispersal limitation as a mechanism for determining microbial community composition. Distance decay relationships also became stronger with increasing distance between facilities. Seasonal variation in community composition was also observed from selected bioreactors, but there was no clear seasonal pattern in the distance decay relationships. IMPORTANCE Understanding the spatiotemporal patterns of biodiversity is a central goal of ecology. The distance decay of community similarity is one of the spatial scaling patterns observed in many forms of life, including plants, animals, and microbial communities. Municipal wastewater treatment relies on microorganisms to prevent the release of excessive quantities of nutrients and other pollutants, but relatively few studies have explored distance decay relationships in wastewater treatment bioreactors. Our results demonstrate a strong distance decay pattern in wastewater treatment bioreactors, regardless of the sequence clustering method or the community dissimilarity metric. Our results suggest that microbial communities in wastewater treatment bioreactors are not randomly assembled but rather exhibit a statistically significant spatial pattern.
Project description:Bioaugmentation of bioreactors focuses on the removal of xenobiotics, with little attention typically paid to the recovery of disrupted reactor functions such as ammonium-nitrogen removal. Chloroanilines are widely used in industry as a precursor to a variety of products and are occasionally released into wastewater streams. This work evaluated the effects on activated-sludge reactor functions of a 3-chloroaniline (3-CA) pulse and bioaugmentation by inoculation with the 3-CA-degrading strain Comamonas testosteroni I2 gfp. Changes in functions such as nitrification, carbon removal, and sludge compaction were studied in relation to the sludge community structure, in particular the nitrifying populations. Denaturing gradient gel electrophoresis (DGGE), real-time PCR, and fluorescent in situ hybridization (FISH) were used to characterize and enumerate the ammonia-oxidizing microbial community immediately after a 3-CA shock load. Two days after the 3-CA shock, ammonium accumulated, and the nitrification activity did not recover over a 12-day period in the nonbioaugmented reactors. In contrast, nitrification in the bioaugmented reactor started to recover on day 4. The DGGE patterns and the FISH and real-time PCR data showed that the ammonia-oxidizing microbial community of the bioaugmented reactor recovered in structure, activity, and abundance, while the number of ribosomes of the ammonia oxidizers in the nonbioaugmented reactor decreased drastically and the community composition changed and did not recover. The settleability of the activated sludge was negatively influenced by the 3-CA addition, with the sludge volume index increasing by a factor of 2.3. Two days after the 3-CA shock in the nonbioaugmented reactor, chemical oxygen demand (COD) removal efficiency decreased by 36% but recovered fully by day 4. In contrast, in the bioaugmented reactor, no decrease of the COD removal efficiency was observed. This study demonstrates that bioaugmentation of wastewater reactors to accelerate the degradation of toxic chlorinated organics such as 3-CA protected the nitrifying bacterial community, thereby allowing faster recovery from toxic shocks.
Project description:Studies investigating the feasibility of new, or improved, biotechnologies, such as wastewater treatment digesters, inevitably start with laboratory-scale trials. However, it is rarely determined whether laboratory-scale results reflect full-scale performance or microbial ecology. The Expanded Granular Sludge Bed (EGSB) bioreactor, which is a high-rate anaerobic digester configuration, was used as a model to address that knowledge gap in this study. Two laboratory-scale idealizations of the EGSB-a one-dimensional and a three- dimensional scale-down of a full-scale design-were built and operated in triplicate under near-identical conditions to a full-scale EGSB. The laboratory-scale bioreactors were seeded using biomass obtained from the full-scale bioreactor, and, spent water from the distillation of whisky from maize was applied as substrate at both scales. Over 70 days, bioreactor performance, microbial ecology, and microbial community physiology were monitored at various depths in the sludge-beds using 16S rRNA gene sequencing (V4 region), specific methanogenic activity (SMA) assays, and a range of physical and chemical monitoring methods. SMA assays indicated dominance of the hydrogenotrophic pathway at full-scale whilst a more balanced activity profile developed during the laboratory-scale trials. At each scale, Methanobacterium was the dominant methanogenic genus present. Bioreactor performance overall was better at laboratory-scale than full-scale. We observed that bioreactor design at laboratory-scale significantly influenced spatial distribution of microbial community physiology and taxonomy in the bioreactor sludge-bed, with 1-D bioreactor types promoting stratification of each. In the 1-D laboratory bioreactors, increased abundance of Firmicutes was associated with both granule position in the sludge bed and increased activity against acetate and ethanol as substrates. We further observed that stratification in the sludge-bed in 1-D laboratory-scale bioreactors was associated with increased richness in the underlying microbial community at species (OTU) level and improved overall performance.
Project description:The effects of two powdered mineral materials (powdered ceramsite and powdered limestone) on aerobic granulation of sludge were evaluated. The experiment was conducted on a laboratory scale bioreactors treating wastewater for 89 days. Three granular sequencing batch reactors (GSBRs) were operated at the lowest optimal organic loading rate (OLR) of 2.55 g COD/(L?d). In the control reactor (R1), the mean diameter (d) of the biomass ranged from 124.0 to 210.0 µm, and complete granulation was not achieved. However, complete granulation did occur in reactors to which either ceramsite (251.9 µm < d < 783.1 µm) or limestone (246.0 µm < d < 518.9 µm) was added. Both powdered materials served as a ballast for the sludge flocs making up the seed sludge. Ceramsite particles also acted as microcarriers of granule-forming biomass. The granules in the reactors with added powdered materials had nonfibrous and smoother surfaces. The reactor with ceramsite exhibited the highest average efficiencies for COD, total nitrogen, and total phosphorus removal (85.4 ± 5.4%, 56.6 ± 10.2%, and 56.8 ± 9.9%, respectively). By contrast, the average nitrification efficiency was 95.1 ± 12.8%.
Project description:Although membrane fouling is a major issue when operating membrane bioreactors (MBRs), information regarding MBR performance and the sludge microbiome after the development of fouling remains limited. For the present study, two MBRs were operated for approximately 1 month under conditions of membrane fouling to investigate the effects of highly stressed environments on the sludge microbiome. After the development of fouling, a Collimonas-related operational taxonomic unit (OTU) was highly dominant in both reactors (relative abundances were ? 63%) and this predomination caused a precipitous decline in the diversity indices of the sludge microbiomes. Because the excessive predomination by limited numbers of OTUs can lead to reductions in the adaptability to environmental changes, monitoring microbial diversity may be a valuable indicator for maintaining the robustness of a sludge microbiome. While, the decrease in the abundance of the Collimonas-related OTU resulted in the predomination of distinct microorganisms in each of the reactors despite being operated under the same conditions; this finding indicates existence of strong pressure to perturb the microbiomes. Detailed analyses suggested that the availability of terminal electron acceptors and competitive interactions between microbes via the secretion of extracellular proteins appeared to differentiate the structures of the respective microbial communities. During the extracellular proteins were secreted in the sludge, considerable portion of microbes were dead and large amounts of biomolecules seemed to be released; resultantly facilitated the predomination of fermentative anaerobes in one reactor as they use organic substances but not inorganic terminal electron acceptors to generate ATP under anaerobic conditions.